Analogues of pyrrolo[3,2d]pyrimidin-4-ones

ABSTRACT

The present invention is novel derivatives of pyrrolo[3,2-d]pyrimidines and pharmaceutical compositions and methods of use therefor. The derivatives are inhibitors of purine nucleoside phosphorylase selectively cytotoxic to T-cells but not to B-cells in the presence of 2&#39;-deoxyguanosine and, therefore, are for use in the treatment of autoimmune diseases, gout, psoriasis or rejection of transplantation.

This is a continuation of U.S. application Ser. No. 117,352, filed Nov.12, 1987, now abandoned, which is a continuation-in-part of U.S.application Ser. No. 059,419 filed June 18, 1987, now abandoned which isa continuation in part of U.S. application Ser. No. 900,486 filed Aug.26, 1986 now abandoned.

BACKGROUND OF THE INVENTION

Various purine derivatives are known including guanine derivativeshaving activity as inhibitors of purine nucleoside phosphoxylase (PNP-4)Ser. No. 767,202 filed Aug. 22, 1985, which is a continuation of U.S.Ser. No. 660,152 filed Oct. 12, 1984, now abandoned. Selected guaninederivatives previously known are also disclosed in the application.Therefore, U.S. Ser. No. 767,202, now U.S. Pat. No. 4,772,606 isincorporated herein by reference.

More specifically the following pyrrolo[3,2-d]pyrimidin-4-ones havingthe Formula ##STR1## are known. The compound of Formula 1 is disclosedin J. Org. Chem., 1979, 44:3826 and the compounds of Formula 2, 3, and 4are disclosed by J. d. Stoeckler, et al., Cancer Res., 1986, 46:1774).Further, Stoeckler, et al disclose PNP activity for compounds of theFormula 2, 3, and 4. These compounds differ from the present inventionby an aryl or heteroaryl substituent compared to the methyl in thecompound of Formulae 1 and the sugar moiety in the compounds of Formula2, 3, and 4. Thus, the present invention are compounds which are notobvious variants thereof.

SUMMARY OF THE INVENTION

The present invention relates to a compound of the Formula (I) ##STR2##Wherein R₆ is OH or SH; R₂ is hydrogen or NH₂, R₈ is hydrogen or NH₂, nis an integer of zero through four, preferably one, and Ar is (i) phenylunsubstituted or substituted by halogen, alkyl of from one to fourcarbon atoms, hydroxy, alkoxy of one to four carbon atoms, ortrifluoromethyl; (ii) 2- or 3-thienyl; or (iii) 2- or 3furanyl; or apharmaceutically acceptable acid or base addition salt thereof.

The present invention also includes methods of manufacturing and apharmaceutical composition for treating autoimmune diseases; such asarthritis, systemic lupus erythematosus, inflammatory bowel diseases,juvenile diabetes, myasthenia gravis, multiple sclerosis, gout and goutyarthritis, as well as psoriasis, viral infections, and cancer, orrejection of transplantation, comprising an immunomodulator orantirejection effective amount; such as a cytotoxic to T-cell amount, ofa compound of the Formula I with a pharmaceutically acceptable carrier.Thus, the invention is also a method of treating an autoimmune disease,psoriasis, or rejection of transplantation as listed above comprisingadministering to a host, such as a mammal including a human sufferingfrom an autoimmune disease or psoriasis or transplantation rejectionadvantageously affected by T-cell toxicity of the compounds of thepresent invention comprising administering an effective amount of acompound of the Formula I in unit dosage form. It is understood, anordinarily skilled physician would begin treatment with a nontoxic andless than effective amount and increase the dose until the desiredeffect is obtained exercising care to administer an amount less than theamount toxic to the host of the disease.

The present invention also includes novel intermediates as follows:

(1) A compound of the Formula (II) ##STR3## Wherein R₂ and Ar is asdefined above; and

(2) A compound of the Formula (III) ##STR4## Wherein R₂ and Ar is asdefined above.

The novel processes of the present invention is as follows:

(A) A process for the preparation of a compound of the Formula Icomprising the treatment of a compound of the Formula II as definedabove with Na₂ S₂ O₄ and then a basic solution such as 1N NaOH in thepresence of heat to obtain the compound of Formula I wherein R₈ isamino. Alternatively the nitro compound II as defined above can bereduced catalytically to give I and overreduction of II in the presenceof a catalyst and hydrogen to obtain a compound of the Formula I whereinR₈ is hydrogen, and, if desired, where R₆ is O or OH, converting saidcompound to a compound where R₆ is S or SH by methods analogous to thoseknown in the art. Such conversion may be performed on the compound ofeither Formula I wherein R₈ is NH₂ or Formula I wherein R₈ is H.Finally, the compound of Formula I may be used to prepare apharmaceutically acceptable acid addition or base salt thereof.

(B) A process for the preparation of a compound of the Formula II asdefined above comprising the treatment of a compound of the Formula IIIas defined above with (1) a base, such as a 1N NaOH solution at aboutroom temperature and then (2) an acid, such as hydrochloric acid toobtain the compound of Formula II.

(C) A process for the preparation of a compound of the formula III asdefined above which comprises contacting a compound of the Formula IIIas defined above which comprises contacting a compound of the Formula(IV) ##STR5## Wherein R₂ is as defined above; with a compound of theformula (V) ##STR6## where in Ar is as defined above; in the presence ofsodium hydride in dimethylformamide or potassium t-butoxide in DMSO orsimilar conditions.

Under certain circumstances it may be necessary to protect either the Nor O of intermediates in the above noted process with suitableprotecting groups which are known. Introduction and removal of suchsuitable oxygen and nitrogen protecting groups are well known in the artof organic chemistry; see for example, (1) "Protective Groups in OrganicChemistry," J. F. W. McOmie, ed., (New York, 1973), pp 43ff, 95ff; (2)J. F. W. McOmie, Advances in Organic Chemistry, Vol. 3, 191-281 (1963);(3) R. A. Borssonas, Advances in Organic Chemistry, Vol. 3, 159-190(1963); and (4) J. F. W. McOmie, Chem. & Ind., 603 (1979).

Examples of suitable oxygen protecting groups are benzyl,t-butyldimethylsilyl, methyl, isopropyl, ethyl, tertiary butyl,ethoxyethyl, and the like. Protection of an N--H containing moiety isnecessary for some of the processes described herein for the preparationof compounds of this invention. Suitable nitrogen protecting groups arebenzyl, triphenylmethyl, trialkylsilyl, trichloroethylcarbamate,trichloroethyoxycarbonyl, vinyloxycarbamate, and the like.

Under certain circumstances it is necessary to protect two differentoxygens with dissimilar protecting groups such that one can beselectively removed while leaving the other in place. The benzyl andt-butyldimethylsilyl groups are used in this way; either is removable inthe presence of the other, benzyl being removed by catalytichydrogenolysis, and t-butyldimethylsilyl being removed by reaction with,for example, tetra-n-butylammonium fluoride.

In the process described herein for the preparation of compounds of thisinvention the requirements for protective groups are generally wellrecognized by one skilled in the art of organic chemistry, andaccordingly the use of appropriate protecting groups is necessarilyimplied by the processes of the charts herein, although not expresslyillustrated.

The products of the reactions described herein are isolated byconventional means such as extraction, distillation, chromatography, andthe like.

The salts of compounds of Formula I described above are prepared byreacting the appropriate base with a stoichiometric equivalent of theacid compounds of Formula I to obtain pharmacologically acceptable saltsthereof.

The compounds of this invention may also exist in hydrated or solvatedforms.

The novel processes in steps beginning with (C) and continuing through(B) and (A) above may be conducted in a one part process. However, it ispreferred to separate the product of Formula III and the product ofFormula II before proceeding to the next process step.

DETAILED DESCRIPTION

The compounds of Formula I and intermediates of Formula II and III ofthe present invention exist in tautomeric forms as purines or guaninesas illustrated below. Both forms are included as part of the inventionand are indiscriminately described in the specification. ##STR7##

The term "alkyl of one to four carbon atoms" means a straight orbranched hydrocarbon chain up to four carbon atoms such as, for example,methyl, ethyl, propyl, isopropyl butyl, isobutyl, secondary butyl ortertiary butyl. "Alkoxy of one to four carbon atoms includes methoxy,ethoxy, propoxy, butoxy and isomers thereof. Halogen is fluorine,chlorine, bromine, or iodine.

The compounds of Formula I are useful both in the free base form, in theform of base salts where possible, and in the form of acid additionsalts. The three forms are within the scope of the invention. Inpractice, use of the salt form amounts to use of the base form.Appropriate pharmaceutically acceptable salts within the scope of theinvention are those derived from mineral acids such as hydrochloric acidand sulfuric acid, and organic acids such as methanesulfonic acid,benzenesulfonic acid, p-toluenesulfonic acid, isothienic acid, and thelike, giving the hydrochloride, sulfate, methanesulfonate,benzenesulfonate, p-toluenesulfonate, and the like, respectively orthose derived from bases such as suitable organic and inorganic bases.Examples of suitable inorganic bases for the formation of salts ofcompounds of this invention include the hydroxides, of ammonia, sodium,lithium, potassium, calcium, magnesium, aluminum, zinc, and the like.

Salts may also be formed with suitable organic bases. Bases suitable forthe formation of pharmaceutically acceptable base addition salts withcompounds of the present invention include organic bases which arenontoxic and strong enough to form such salts. These organic bases forma class whose limits are readily understood by those skilled in the art.Merely for purposes of illustration, the class may be said to includemono-, di-, and trialkylamines, such as methylamine, dimethylamine, andtriethylamine; mono-, di-, or trihydroxyalkylamines such as mono-, di-,and triethanolamine; amino acids such as arginine, and lysine;guanidine; N-methylglucamine; L-glutamine; N-methylpiperazine;morpholine; ethylenediamine; N-benzylphenethylamine;tris(hydroxymethyl)aminomethane; and the like. (See for example,"Pharmaceutical Salts," J. Pharm. Sci. (1977) 66(1):1-19.)

The acid addition salts of said basic compounds are prepared either bydissolving the free base of compound I in aqueous or aqueous alcoholsolution or other suitable solvents containing the appropriate acid orbase and isolating the salt by evaporating the solution, or by reactingthe free base of compound I with an acid as well as reacting compound Ihaving an acid group thereon with a base such that the reactions are inan organic solvent, in which case the salt separates directly or can beobtained by concentration of the solution.

A preferred embodiment of the present invention is a compound of FormulaI wherein R₆ is OH or SH; R₂ and R₈ are NH₂, n is one, and Ar is eitherphenyl or 2- or 3-thienyl. A more preferred embodiment is2,6-diamino-3,5-dihydro-7-(phenylmethyl)-4H-pyrrolo[3,2-d]pyrimidin4-one,2,6-diamino-3,5-dihydro-7-(2-thienylmethyl)-4H-pyrrolo[3,2-d]pyrimidin-4-oneor2,6-diamino-3,5-dihydro-7-(3-thienylmethyl)-4H-pyrrolo[3,2-d]pyrimidin4-one.Of these the most preferred is the latter of these three.

The compound of Formula I may generally be prepared according to themethod shown in the following scheme. ##STR8##

Generally, the starting materials having the Formula IV and V are eithercommercially available or may be prepared by known methods. For example,the compounds of Formula IV may be prepared by a procedure described inSynthetic Procedures in Nucleic Acid Chemistry Vol. 1 (1973) page 94,and the compounds of Formula V may be prepared by procedures analogousto those described in U.S. Pat. No. 4,279,903 and references citedtherein.

Further, the conditions for contacting the compounds V and IV of process(C) above include the dropwise addition of sodium hydride oil suspensionto a solution of the compound V in a solvent such as dimethylformamide(DMF) followed by the further addition of a solution of the compound offormula IV also in a solvent such as DMF under a dry nitrogenatmosphere. The resulting reaction mixture is heated to from 60° to 90°C. for from 30 minutes to two hours and then acidified to about pH 3 or4 with 1-6 N HCl. The condensation of process (c) may also beaccomplished in a solvent, such as DMSO, and in the presence oft-butoxide.

The conditions of process (B) for treating the product of (C) having theFormula III as defined above are first treating a solution thereof with,for example, 1N NaOH at about room temperature followed by acidifictionwith, for example 4N HCl to precipitate the product having the FormulaII.

For process (A) a solution of the compound of Formula II in 1N NaOH istreated with sodium dithionite at from 70° to 95° C. and preferably 90°C. for from 10 to 60 minutes, preferably about 25 to 40 minutes and thenagain acidified to about pH 3 or 4 to provide crystals of the product ofFormula I wherein R₆ is oxygen and R₈ is NH₂. Further catalyticreduction under conditions within those known to the artisan for similarreductions the Compound II wherein R₆ is oxygen and R₈ is amino orhydrogen is obtained.

The compounds having the Formula I of the present invention have beenshown to exhibit significant enzyme inhibition activity and cytotoxicactivity. In the purine nucleoside phosphorylase (PNP-4) enzyme assay,an IC₅₀ is achieved at a dose of about from 0.9 to 3.18 micromoles onselected compounds of the present invention. PNP-4 activity for thecompounds of Formula I is measured radiochemically by measuring theformation of [¹⁴⁻ C]-hypoxanthine from [¹⁴⁻ C]inosine [Biomedicine, 33,39 (1980)] using human erythrocyte as the enzyme source. An in vivoinhibition of purine nucleoside phophorylase (HPLC-1) enzyme assay isused essentially as disclosed in the Annals of New York Academy ofSciences, Volume 451, Page 313 (1985) to further show the activity ofthe compounds of Formula I of the present invention. The same compoundsalso were found by a standard test (HTBA-1) [Science, 214, 1137, (1981)]to be selectively cytotoxic for T-cells in the presence of2'-deoxyguanosine at a similar concentration range and nontoxic toB-cell in the presence of the same amount of 2'-deoxyguanosinedemonstrating utility for the compounds of Formula I as describedherein. Since removal of T-cells or modulation of T-cells are beneficialin the treatment of autoimmune diseases, these compounds beingselectively cytotoxic to T-cells will, therefore, be useful in theirtreatment. 8-aminoguanosine, a known PNP-inhibitor, has been shown to beefficacious for inhibiting rejection of skin graft in dogs [J. B.Benear, et al, Transplantation, 1986, 41:274]. Clinically it has beenshown that modulation and/or removal of T-cells by throacic ductdrainage, lymphapheresis or total lymphoid irradiation gave partial tocomplete relief from rheumatoid arthritis in patients who were totallyrefractory to other forms of therapy (A. Tanay, et al, Arthritis andRheumatism, Vol. 30, No. 1, p 1 (1987). S. Strober, et al, Annual ofInternal Medicine, V-102, No. 4, 441-449 (1985); H. G. Nusslein, et al,Arthritis and Rheumatism, V-28, No. 11, 1205-1210 (1985); E. Brahn, etal, ibid, V-27, No. 5, 481-487 (1984), and J. Karsh, et al, ibid, V-24,No 7, 867-873 (1981)). Cyclosporin A, a T-cell modulator, showedbeneficial effects in the treatment of juvenile diabetes. (A. Assan, etal, The Lancet, January 12, p 67, (1985). Additionally, cyclosporin A ispresently the drug of choice for the prevention of transplant rejection,(R. M. Merion, et al, New Eng. J. Med., (1984), 148). More recently,cyclosporin A is shown to be useful to treat psoriasis. Further, it issuggested the cyclosporin therapy is shown to markedly reduce activatedT-cells in psoriatic lesions. Therefore, it is reasonable to believe thebasis of the successful treatment of psoriasis is modulation of T-cellactivity. (See C. N. Ellis, et al, JAMA. V-256, No. 22, Dec. 12, 1986,pp 3110-3116.) Finally, cyclosporin A is shown to be efficacious inrheumatoid arthritis. (M. E. Weinblatt, et al, Arthritis and Rheumatism,V-30, No. 1 pp 11-17 (January, 1987); O. Forre, et al, Arthritis andRheumatism, V-30, No. 1, pp 88-92 (January, 1987); M. Dougados, et al,Arthritis and Rheumatism, Vol. 30, No. 1, pp 83-87 (January, 1987);

Representative examples from the prsent invention are shown in thefollowing activity table to provide the activity discussed above.

                  ACTIVITY TABLE                                                  ______________________________________                                         ##STR9##                                                                                      HTBA-1                                                                        T-cell +  HPLC-1                                                        PNP-4 10 μM  (mg/kg; PO)                                        Num-                 IC.sub.50                                                                           2'-d Gua;                                                                             Inosine                                    ber   R.sub.8                                                                              Ar*     (μM)                                                                             IC.sub.50 μM                                                                       (μM)                                                                             Guanosine                            ______________________________________                                        3     NH.sub.2                                                                             C.sub.6 H.sub.5                                                                       1.6   1.7     2.93  0.54 (100)                           6     NH.sub.2                                                                             2-Th    1.0   1.9     2.64  0.36 (100)                           9     NH.sub.2                                                                             3-Th    0.9   2.3     2.62  0.26 (150)                           13    NH.sub.2                                                                             2-meth  3.18 or                                                               oxy-    3.09                                                                  phenyl                                                           ______________________________________                                         Th = Thiophene                                                           

In vivo studies based on the above noted disclosures maybe used todetermine activity in the particular disease states noted.

Since T-Cells play a central role in immune response, use of thecompounds of the invention is contemplated for the immunoregulation toprevent rejection in transplantation or in the treatment of psoriasisand in the treatment of autoimmune disease such as rheumatoid arthritis,systemic lupus erythrematosus, inflammatory bowel disease, multiplesclerosis, myasthemia gravis, gout or gouty arthritis juvenile diabetes,cancer, and viral diseases. The present invention thus includescompositions containing a compound of Formula I in treating rejection oftransplantation or disease such as psoriasis in humans or autoimmunedisease characterized by abnormal immune response in primates or humans.According to this aspect of the invention, the properties of thecompounds of the invention are utilized by administering to awarmblooded animal an effective amount of a pharmaceutical compositioncontaining as the active ingredient at least about 0.1 percent byweight, based on the total weight of the composition of at least onesuch compound of the invention.

Pharmaceutical compositions of the invention can be formulated in anysuitable way, preferably with an inert carrier for administrationorally, parenterally, ophthalmically, topically, or by suppository.

For example, the compounds of the present invention are formulated intodosage forms such as tablets or syrups by blending with an inertpharmaceutical carrier such as lactose or simple syrup by methods wellknown in the art. For injectable dosage forms, they are formulated withvehicles such as water, propylene glycol, peanut oil, sesame oil, andthe like. In these dosage forms, the active ingredient is from about0.05 grams to 0.5 grams per dosage unit.

The present invention is further illustrated by way of the followingexamples. ##STR10##

EXAMPLE 12-Amino-α-cyano-1,6-dihydro-5-nitro-α-(phenylmethyl)-6-oxo-4-pyrimidineaceticacid, ethyl ester

A solution of ethyl 2-cyano-3-phenyl-propionate (JCS, 1944, 13) (6.0 g)in DMF (25 mL) is added dropwise to a suspension of sodium hydride (1.2g, 60% suspension in oil, washed with hexane) in DMF (25 mL) under anatomosphere of dry N₂. The reaction mixture is stirred for 15 min atroom temperature and then a solution of2-amino-6-chloro-5-nitro-4(3H)-pyrimidinone (Synthetic Procedures inNucleic Acid, V-1, 1973, p. 94) (1.9 g, freshly crystallized frommethanol) in DMF (50 mL) is added to the anion. The reaction mixture isheated at 65° C. for 24 h and then acidified to pH 3 with 1N HCl.

Most of the DMF is removed by evaporation on a rotary evaporator underhigh vacuum. The residual oil is partitioned between ethyl acetate (500mL) and water. The organic layer is dried (Na₂ SO₄) evaporated to agummy solid. The residue is triturated with ether and collected byfiltration. It is dried under vacuum to give2-Amino-α-cyano-1,6-dihydro-5-nitro-α-(phenylmethyl)-6-oxo-4-pyrimidineaceticacid, ethyl ester as a partial hydrate (2.25 g, 62%). ##STR11##

EXAMPLE 22-Amino-1,6-dihydro-5-nitro-6-oxo-α-(phenylmethyl)-4-pyrimidineacetonitrile

A solution of2-Amino-α-cyano1,6-dihydro-5-nitro-α-(phenylmethyl)-6-oxo-4-pyrimidineaceticacid, ethyl ester as prepared above (2.0 g) in 1N NaOH (100 mL) isstirred at room temperature for 1 h and is acidified with 4N HCl (30mL). The precipitate is collected by filtration and dried under vacuumto give2-Amino-l,6-dihydro-5-nitro-6-oxo-α-(phenylmethyl)-4-pyrimidineacetonitrile(600 mg, 36%). ##STR12##

EXAMPLE 32,6-Diamino-3,5-dihydro-7-(phenylmethyl)-4H-pyrrolo[3,2-d]pyrimidin-4-one

To a solution of2-Amino-1,6-dihydro-5-nitro-6-oxo-α-(phenylmethyl)-4-pyrimidineacetonitrile as prepared above (10.0 g) in 1N NaOH (600 mL) is addedsodium dithionite (35 g). The reaction mixture is heated at 90° C. for35 min and is acidified to pH 4 with 4N HCl while still hot. Thereaction mixture is cooled in an ice bath and the precipitate iscollected by filtration. It is dried over P₂ O₂ under vacuum (4.0 g).The crude (undecarboxy-ated) product is dissolved in 300 mL conc HCl andquickly filtered through a glass frit before it crystallized out. Theresulting suspension is boiled for five minutes and cooled. The product2,6-Diamino-3,5-dihydro-7-(phenylmethyl)-4H-pyrrolo[3,2-d]pyrimidin-4-one is collectedby filtration (1.98 g, 23%) mp >250° C. (dec). ##STR13##

EXAMPLE 42-Amino-α-cyano-1,6-dihydro-5-nitro-6-oxo-α-(2-thienylmethyl)-4-pyrimidineacetic acid, methyl ester

Sodium hydride (4.5 g, 60% suspension in oil washed with hexane) issuspended in dry DMF (50 mL) under an atmosphere of dry N₂ and asolution of methyl 2-cyano- 3-(2-thienyl) propionate (U.S. Pat. No.4,279,903) (22.0 g) in dry DMF (50 mL) is added dropwise when a darkblue solution is formed. A solution of freshly recrystallized2-amino-6-chloro-5-nitro-4(3H)-pyrimidinone (8.57 g) in DMF (75 mL) isadded in one portion. The reaction mixture is heated at 60° C. overnightand cooled and then is acidified to pH 5 with 1N HCl. It is poured into1000 mL ethyl acetate and is extracted with water (4×300 mL). Theorganic layer is evaporated to near dryness and the residue is suspendedin ether and collected by filtration. The crude product is washed withhexane until the washings are no longer green. The solid is dried undervacuum to give2-Amino-α-cyano-1,6-dihydro-5-nitro-6-oxo-α-(2-thienylmethyl)-4-pyrimidine-aceticacid, methyl ester. ##STR14##

EXAMPLE 52-Amino-1,6-dihydro-5-nitro-6-oxo-α-(2-thienylmethyl)-4-pyrimidineacetonitrile

A solution of2-Amino-α-cyano-1,6-dihydro-5-nitro-6-oxo-α-(2-thienylmethyl)-4-pyrimidineaceticacid, ethyl ester as prepared above (5.0 g) in 1N NaOH (200 mL) isstirred at room temperature for 90 min. The reaction mixture isacidified to pH 1 with 4N HCl and stirred for 5 min. The reactionmixture is neutralized (pH 7) with 1N NaOH and the product2-Amino-1,6-dihydro-5-nitro-6-oxo-α-(2-thienylmethyl)-4-pyrimidineacetonitrile(4.1 g, 98%) is collected by filtration. ##STR15##

EXAMPLE 6 2,6-Diamino-3,5-dihydro-7-(2-thienylmethyl)-4H-pyrrolo[3,2-d]pyrimidin-4-one

To a solution of2-Amino-1,6-dihydro-5-nitro-6-oxo-α-(2-thienylmethyl)-4-pyrimidineacetonitrileas prepared above (4.0 g) in 1N NaOH (250 mL) is added sodium dithionite(17 g) and the reaction mixture is heated at 90° C. for 20 min. Thereaction mixture is acidified to pH 2 with 4N HCl while still hot andfiltered and then is cooled and neutralized with 1N NaOH. The resultingprecipitate is collected by filtration and dried. The crude product isadded to a stirred solution of conc HCl (150 mL) and the hydro-chloridesalt is collected by filtration. The salt is dissolved in aq NaOH andreprecipitated with 1N HCl. Recrystallization from 2N HCl gave theanalytical hydrochloride salt the monohydrate, of2,6-Diamino-3,5-dihydro-7-(2-thienylmethyl)-4H-pyrrolo[3,2-d]-pyrimidin-4-one (0.82, 19%), mp 220°-225° C. (dec).##STR16##

EXAMPLE 72-Amino-α-cyano-1,6-dihydro-5-nitro-6-oxo-α-(3-thienylmethyl)-4-pyrimidineaceticacid, ethyl ester

Sodium hydride (5.7 g; 60% suspension in oil is was with hexane) issuspended in dry DMF (50 mL) under an atmosphere of dry N₂ and asolution of methyl 2-cyano-3-(3-thienyl) propionate (U.S. Pat. No.4,279,903) (28.0 g) in DMF (50 mL) is added dropwise.2-Amino-6-chloro-5-nitro-4 (3H)-pyrimidinone (9.1 g, freshlyrecrystallized) in DMF (50 mL) is added. The reaction mixture is heatedat 70° C. overnight, cooled, and then acidified to pH 4 with 1N HCl withice bath cooling. The reaction mixture is diluted to 1000 mL with coldwater and the resulting precipitate is collected by filtration. It isrinsed with hexane/ethylacetate and dried to give2-Amino-α-cyano-1,6-dihydro-5-nitro-6-oxo-α-(3-thienylmethyl)-4-pyrimidine-aceticacid, ethyl ester (Ar=3-Th) (8.8 g, 52%). ##STR17##

EXAMPLE 82-Amino-1,6-dihydro-5-nitro-6-oxo-α-(3-thienylmethyl)-4-pyrimidineacetonitrile

A solution of2-Amino-α-cyano-1,6-dihydro-5-nitro-6-oxo-α-(3-thienyl-methyl)-4-pyrimidineaceticacid, ethyl ester in 1N NaOH (200 mL) is stirred for 2 h at roomtemperature and then acidified to pH 1 by the dropwise addition of concHCl. The resulting suspension is warmed (45° C.) for 2 min and thencooled. The pH is adjusted to pH=3 with NH₄ OH. The solid is collectedby filtration, washed with water, and dried under vacuum to give2-Amino-l,6-dihydro-5-nitro-6-oxo-α-(3-thienylmethyl)-4-pyrimidineacetonitrile(3.03 g). ##STR18##

EXAMPLE 92,6-Diamino-3,5-dihydro-7-(3-thienylmethyl)-4H-pyrrolo[3,2-d]pyrimidin-4-on

To a solution of2-Amino-1,6-dihydro-5-nitro-6-oxo-α-(3-thienylmethyl)-4-pyrimidineacetonitrileprepared above (5.0 g) in 1N NaOH (300 mL) is added sodium dithionite(20 g). The reaction mixture is heated for 30 min at 90° C. and then isacidified (pH 1) with conc HCl while still hot. The reaction mixture iscooled and neutralized with ammonium hydroxide. The resultingprecipitate is collected by filtration, washed with cold water and driedunder vacuum. The crude product is added to 100 mL conc HCl in smallportions with stirring and the hydrochloride salt is collected byfiltration. The product is recrystallized from 1N HCl to give2,6-Diamino-3,5-dihydro-7-(3-thienylmethyl)-4H-pyrrolo[3,2-d]pyrimidin-4-one as the monohydro-chloride salt (2.35 g), mp>185°C. (dec). ##STR19##

EXAMPLE 102-Amino-α-cyano-α-(2-furanylmethyl)-1,6-dihydro-5-nitro-6-oxo-4-pyrimidineaceticacid, ethyl ester

Sodium hydride (8.4 g, 60% suspension in oil, washed with hexane) issuspended in dry DMF (100 mL) under an atmosphere of dry N₂ and asolution of methyl 2-cyano-3-(2-furanyl)propionate (U.S. Pat. No.4,279,903) (37.6 g) in DMF (100 mL) is added dropwise. When the additionis complete, a clear red solution is formed.2-Amino-6-chloro-5-nitro-4(3H)-pyrimidinone (recrystallized frommethanol) (13.34 g) is added as a solid. The reaction mixture is heatedat 100° C. for 1 h and at 65° C. overnight. The reaction mixture iscooled in an ice bath, acidified with 10% aq HCl and diluted with enoughwater to precipitate out the product2-Amino-α-cyano-α-(2-furanylmethyl)-1,6-dihydro-5-nitro-6-oxo-4-pyrimidineaceticacid, ethyl ester. The product is collected by filtration and rinsedwith water and ether (18.5 g). ##STR20##

EXAMPLE 112-Amino-α-(2-Furanylmethyl)-1,6-dihydro-5-nitro-6-oxo-4-pyrimidineacetonitrile

A solution of2-Amino-α-cyano-α-(2-furanyl-methyl)-1,6-dihydro-5-nitro-6-oxo-4-pyrimidineaceticacid, ethyl ester (5.0 g) in 1N NaOH (200 mL) is stirred at roomtemperature for 2 h. The reaction mixture is acidified (pH 1) by thedropwise addition of conc HCl and is stirred at room temperature for 2min. The precipitate is collected by filtration, washed with water, anddried to give2-Amino-α-(2-Furanylmethyl)-1,6-dihydro-5-nitro-6-oxo-4-pyrimidineacetonitrile(3.02 g 73%). ##STR21##

EXAMPLE 12 2,6-Diamino-3,5-dihydro-7-(2-furanylmethyl)-4H-pyrrolo[3,2-d]pyrimidin-4-one

To a solution of2-Amino-α-(2-Furanylmethyl)-1,6-dihydro-5-nitro-6-oxo-4-pyrimidineacetonitrile(4.0 g) in 1N NaOH (250 mL) is added sodium dithionite (16 g) and thereaction mixture is heated at 90° C. for 30 min. The reaction mixture iscooled in an ice bath and neutralized with 4N HCl. The resultingprecipitate is collected by filtration and purified by a series ofacid/base reprecipitations. First the product is reprecipitated from 1NNaOH by acidifying (pH 2) with a saturated solution of oxalic acid. ThisNaOH/oxalic acid reprecipitation is repeated. Then it is reprecipitatedfrom 1N NaOH by adjusting pH to 11 with 4N HCl. The product2,6-Diamino-3,5-dihydro-7-(2-furanylmethyl)-4-4H-pyrrolo[3,2-d]pyrimidin-4-oneis dried under vacuum.

EXAMPLE 13

The following compound is prepared according to the method of Example 3above using appropriate corresponding starting materials:2,6-diamino-3,5-dihydro-7-(2'-methoxyphenylmethyl)-4H-pyrrolo[3,2-d]pyrimidin-4-onemonohydrochloride with a third mole H₂ O mp 215°-240° C. (dec).

We claim:
 1. A compound of the formula (I) ##STR22## wherein R₆ is OH orSH; R₂ is hydrogen or NH₂, R₈ is hydrogen or NH₂, n is an integer of onethrough four, and Ar is (i) phenyl unsubstituted or substituted byhalogen, alkyl of from one to four carbon atoms, hydroxy, alkoxy of oneto four carbon atoms, or trifluoromethyl, (ii) 2- or 3- thienyl, (iii)2- or 3- furanyl; or a pharmaceutically acceptable acid or base additionsalt thereof.
 2. A compound of claim 1 wherein n is one.
 3. A compoundof claim 1 wherein R₆ is SH.
 4. A compound of claim 1 wherein R₆ is OH.5. A compound of claim 3 wherein R₂ and R₈ is NH₂.
 6. A compound ofclaim 4 wherein R₂ and R₈ are NH₂.
 7. A compound of claim 6 and being 2,6-diamino-3,5-dihydro-7-(phenylmethyl)-4H-pyrrolo[3,2-d]pyrimidin-4-one.8. A compound of claim 7 and being the monohydrochloride salt thereof.9. A compound of claim 6 and being 2,6-diamino-3,5-dihydro-7-(2-thienylmethyl)-4H-pyrrolo[3,2-d]pyrimidin-4-one.10. A compound of claim 9 and being the monohydratehydrochloride saltthereof.
 11. A compound of claim 6 and being2,6-diamino-3,5-di-hydro-7-(3-thienylmethyl)-4H-pyrrolo[3,2-d]pyrimidin-4-one.12. A compound of claim 11 and being the monohydrochloride salt thereof.13. A compound of claim 11 and being the organic acid addition salts oraddition salts of amino acids thereof.
 14. A compound of claim 6 andbeing2,6-diamino-3,5-dihydro-7-(2-furanylmethyl)-4H-pyrrolo-[3,2-d]pyrimidin-4-one.15. A compound of claim 6 and being2,6-diamino-3,5-dihydro-7-(2'-methoxyphenylmethyl)-4H-pyrrolo[3,2-d]pyrimidin-4-one.16. A pharmaceutical composition for treating autoimmune diseasesadvantageously affected by selective toxicity to T-cells or rejection oftransplantation comprising a cytotoxic to T-cell amount of a compound offormula I of claim 1 with a pharmaceutically acceptable carrier.